Toxoplasma gondii is an obligate intracellular pathogen with a remarkably broad host range; it is able to infect virtually any nucleated cell of almost any warm-blooded animal. While well-known for the damage Toxoplasma can cause to a developing fetus, it has become a serious opportunistic pathogen in HIV/AIDS and lymphoma patients, as well as in organ transplant patients who are undergoing immunosuppressive therapy. Different strains of Toxoplasma can lead to different disease outcomes, both in humans and in mice. I have identified the secreted Toxoplasma pseudokinase ROP5 as a dominant virulence factor. This K22 award will provide the resources, time, and training to allow me to build an independent research program to determine molecular mechanism of ROP5's profound effect on disease outcome. This work will provide a much-needed interrogation of the interactions between a pathogen and the mammalian immune system, enabling the development of new therapies for toxoplasmosis. I have found that parasites deficient in ROP5 are completely attenuated in their virulence to mice and elicit a stronger pro-inflammatory response during early infection than do wild-type parasites. Furthermore, I have solved the crystal structure of ROP5's pseudokinase domain and found that polymorphisms correlated with strain-specific differences in virulence form a surface near the substrate binding domain. This has led me to hypothesize that ROP5 subverts the host immune system by binding host signaling molecules in its pseudokinase domain, which leads to dysregulation of immune signaling networks. I will test this hypothesis in the following complementary aims: 1) Identify the specific immune cell types and signaling networks modulated by ROP5; 2) Determine the physical interactions ROP5 makes in infected host cells and the biochemical mechanism of ROP5's profound effect on virulence.